Scavenger receptors (SR) are cell surface proteins that bind chemically modified lipoproteins and exhibit broad ligand binding specificities. Members of the class B family include SR-BI (and its alternatively spliced form SR-BII), which is an HDL receptor involved directly or indirectly in many mammalian physiologic and pathophysiologic processes. These include maturation and function of blood cells, female fertility, inflammation, responses to viral and bacterial infection and to apoptosis, movement of lipids into and out of cells, and-most directly-lipoprotein metabolism and its associated diseases (atherosclerosis, coronary heart disease). SR-BI controls plasma HDL levels and delivers HDL's cholesterol to hepatocytes and steroidogenic and other cells via selective lipid uptake, a poorly understood mechanism distinct from classic receptor-mediated endocytic uptake of lipoproteins. SR-BI also mediates cellular cholesterol efflux to lipoproteins. In hepatocytes in vivo SR-BI expression depends on its interaction with a cytoplasmic, scaffolding, adaptor protein called PDZK1. The overall goals of this proposal are 1) to more precisely define how ligands bind to SR-BI, 2) to determine the mechanism(s) of SR-BI-mediated lipid transport (selective uptake and efflux). and 3) to determine how the adapter protein PDZK1 mediates tissue-specific regulation of SR-BI abundance, intracellular localization and activity. The work will rely on the generation and functional analysis of mutant receptors, including those generated using a novel approach called Retrovirus Library Based Activity Dissection ('ReLiBAD'). It will also focus on identification and characterization of small molecule inhibitors and activators of SR-BI (Chemical Genetics), which will be used with several biophysical techniques as probes of SR-BI mechanism. Characterization of these small molecules may possibly help direct the design of potential pharmacologic reagents. Detailed characterization of the structure and distinctive binding properties of SR-BI will provide important tools for the analysis of its mechanism of action and functions. Characterization of the molecular mechanisms underlying SR-BI function may suggest new approaches for the treatment and prevention of at least some of its related pathophysiology (e.g., atherosclerosis, infectious disease, female infertility). PUBLIC HEALTH RELEVANCE: A protein on the surfaces of certain cells, called the HDL (good cholesterol) receptor (technical name: SR-BI) appears to be involved directly or indirectly in many mammalian physiologic and pathophysiologic processes, including transformation of an egg cell into a mature adult, mental function during aging, female fertility, red blood cell stability and function, inflammation, the body's response to infection by bacteria and viruses and to the programmed death, movement of fatty substances within and into and out of the body, movement and function and repair of the cells that line blood vessel walls, protection against oxidative stress and the metabolism of cholesterol and the carriers (called lipoproteins) that move cholesterol around the body (e.g., the good cholesterol HDL and the bad cholesterol LDL), as well as diseases associated with cholesterol (e.g., atherosclerosis, coronary heart disease, heart attacks, heart failure). Thus, defining precisely how SR-BI 'works'in the body and identifying drug-like molecules that alter its ability to work (goals of this proposal) will have significant impact on understanding body function and major diseases, and open the door to developing new drugs to treat the nation's number 1 killer-clogging of the arteries, causing heart attacks and strokes. Indeed, recent disappointments in attempts by the pharmaceutical industry to develop HDL-targeted drugs highlight the importance of developing a deeper and broader understanding of all aspects of HDL metabolism, including those mediated by SR-BI.